Summary: Our lab has analyzed the phenotype, location and functional properties of lymphocytes stimulated to produce antibodies and cytokines when challenged with vaccines, conventional protein-based antigens, and DNA derived from bacteria or viruses. This work includes studies examining the immunogenicity of the envelope glycoprotein of HIV induced by natural infection and in animal models involving retroviral infection or the administration of purified recombinant HIV proteins. Also studied was the effect of adjuvants on the activation of cytokine producing cells, and whether early changes in the cytokine milieu effect subsequent immune responses. In these experiments, cells were obtained from mice and humans infected with the HIV or MAIDS virus, or following immunization with purified recombinant HIV envelope glycoproteins (or other protein antigens). The phenotype of the cells activated by each immunogen, and the magnitude, specificity and in vivo localization of the reactive cells, was examined. This work I) established the cytokine profile of cells located in distinct lymphoid organs, ii) enabled us to monitor the effect of selected adjuvants on the cytokines elicited by protein antigens, and iii) showed that immunizing young animals had long-term effects on the Th profile of their resting T cell pool. An antigen-specific restimulation assay was developed, permitting us to demonstrate that cytokine secreting cells activated by in vivo immunization reacted in a specific manner when re-exposed to the same antigen in vitro. In patients with HIV, we found that disease severity correlated with a loss of both type 1 and type 2 cytokine secreting cells, while long-term non-progressors manifest stable cytokine profiles. We showed that the relative ratio (rather than absolute number) of type 1 : type 2 cytokine secreting cells correlated best with disease progression. Declining CD4 cell count correlated with a preferential loss of such Th1 secreting cells. We are in the process of examining the response of cells from HIV patients to CpG-containing DNA motifs, and the ability of such motifs to induce the production of chemokines capable of controling HIV infection. Most recently, we've been studying how changes to the CpG content of DNA vaccines encoding HIV antigens alters the immunogenicity of the resultant vaccine. Data suggest that adding more CpG motifs, and insuring that they do not lie near "suppressive motifs", positively influences immunogenicity.